1. Field of the Invention
This invention pertains to the field of methods for preparing libraries of purine compounds. For example, the invention relates to methods of preparing 2,6,9-substituted purine compounds on solid phase supports by employing sulfenylpurine intermediates. Also provided are methods for preparing 2,9-substituted purines and O6-aryl and O6-alkyl-substituted purines.
2. Background
The sequencing of the human genome and numerous pathogen genomes has resulted in an explosion of new molecular targets that can be modulated by small molecules. Discovering these modulators and further developing them into new therapeutics presents unprecedented opportunities and challenges for academic and industrial researches. One of the most fruitful applications of combinatorial chemistry has been the design of flexible synthetic schemes that generalize a privileged scaffold from a particular target protein to an entire protein family.
Inhibitors of protein kinases have proven to be invaluable tools in the elucidation of signal transduction networks, as well as promising clinical candidates for the treatment of cancer, cardiovascular disease, inflammatory, and neurological diseases. (McMahon et. al., Current Opinion in Drug Discovery & Development, 1, 131 (1998); Adams et al., Current Opinion in Drug Discovery & Development, 2, 96 (1999); Cohen, P., Current Opinion in Chemical Biology, 3, 459 (1999); Garcia-Echeverria et. al., Med. Res. Rev, 20, 28 (2000); Blume-Jensen et al., Nature, 411, 355 (2001) and references therein) Despite concerns that it would be extremely difficult to design specific ATP competitive inhibitors of kinases, there have been a number of success stories including the p38 Map kinase, tyrosine kinases, and cyclin dependent kinases. (McMahon et. al., Current Opinion in Drug Discovery & Development, 1, 131 (1998); Adams et al., Current Opinion in Drug Discovery & Development, 2, 96 (1999); Cohen, P., Current Opinion in Chemical Biology, 3, 459 (1999); Garcia-Echeverria et. al., Med. Res. Rev, 20, 28 (2000); Blume-Jensen et al., Nature, 411, 355 (2001) and references therein; Druker et. al., Nature Medicine, 2, 561 (1996); Taylor et al., Current Opinion in Chemical Biology, 1, 2219 (1997); Schindler et. al., Science, 289, 1938 (2000)) Selective inhibitors of each of these kinases are in various stages of clinical testing. The ability to discriminate between extremely homologous kinases such as CDK1 vs. CDK2 has been demonstrated by the development of novel thio-flavopiridol derivatives that display enhanced selectivity for CDK1 relative to CDK2. (Kim et. al., J. Med. Chem., 43, 4126 (2000).)
The purine ring system is a key structural element of the substrates and ligands of many biosynthetic, regulatory and signal transduction proteins including cellular kinases, G proteins and polymerases. In particular, inhibitors of protein kinases have proven to be invaluable tools in the elucidation of signal transduction networks as well as promising clinical candidates in a multitude of disease such as cancer, cardiovascular disease, inflammatory disease and neurological disease. For example, various purine analogs have been found to be highly potent therapeutic agents for disease. As such, the purine ring system has been a good starting point in the search for inhibitors of kinases, G proteins and many biomedically significant processes.
The design of synthetic schemes for generating a multitude of structurally diverse compounds (i.e., library of compounds) typically involves creating the libraries in situ in solution phase or on solid support, i.e., solid phase. In solid phase synthesis, the desired compounds are generated while attached to the solid support via a linker prepared on a polymeric solid support material, e.g., polystyrene.
Several solid phase and solution phase approaches for the synthesis of purine analogs have been reported in the literature over the past five years. (For 2-, 6-, 8- 9-substituted purine analogs, see, e.g., Gray et. al., Science, 281, 533 (1998); Chang et. al., Chemistry and Biology, 6, 361 (1999); Lucrezia et. al., J. Comb. Chem., 2, 249 (2000); Nolsoe et. al., Synth. Commun., 28, 4303 (1998); for 7-substituted purine analogs, see, e.g., Dalby et. al., Angew. Chem. Int. Ed. Engl., 32, 1696 (1993); Zaitseva et. al., Bioorganic Med. Lett., 5, 2999 (1995).) Unfortunately, these approaches have limitations. One limitation is that one substituent is held invariant in order to anchor the purine ring to the solid phase (Scheme 1). To avoid this limitation, a “traceless” strategy would be desirable that would be compatible with production scale library synthesis in spatially separate or divide-recombine formats. 
Another limitation of previous synthetic approaches is the low reactivity of the 2-fluoro group once an amino substituent has been installed at C6. For example, complete displacement at C2 of a 2-fluoro-6-benzylaminopurine in solution requires heating at over 100° C. for twelve hours using n-butanol as solvent. Complete aromatic substitution of 2-fluoro or 2-chloro purine compounds on solid support requires even higher temperatures and often results in significant side reactions. This limits the range of functional groups that can be installed at C2 and also creates difficulties in library production.
Subsequently, it was found that 6-amino-2-fluoro-9-alkylpurines react with primary amines in methanol at room temperature. With slightly more forcing conditions (in refluxing methanol) sterically hindered amines such as the alpha-amino group of arginine could be successfully introduced at the C2 position (Scheme 2) with good yields. Unfortunately, these conditions failed to translate to solid support, presumably due to resin swelling problems in methanol. Despite testing a range of solvent systems (NMP, DMF, dioxane, DMSO, THF and their combinations such as DMFv/MeOHv:1/1), no solvent was found that allowed complete substitution below 100° C. 
Since a number of purine compounds have been shown to possess diverse pharmacological properties and biological activities in a number of therapeutic areas, the generation of a substituted purine compound library would be useful as a screening tool to identify the structures of compounds that possess the desired biological activity. In addition, the generation of substituted purine library would permit alteration of the structures of the identified compounds to identify derivatives of these compounds which exhibit the best biological activity and which also have ideal pharmacological and pharmacokinetic properties. Accordingly, the development of an efficient, rapid in situ method for the generation of a multitude of highly substituted purine compounds that would overcome limitations of known procedures would be highly desirable.
Moreover, to date a variety of heterocyclic scaffolds including pyrimidines, indolines, pyrrolopyrimidines, indirubins, purines, quinazolines, trisubstituted imidazoles, pyrazolopyrimidines, have been developed as kinase inhibitors. ((a) McMahon et. al., Current Opinion in Drug Discovery & Development, 1, 131 (1998); (b) Adams et al., Current Opinion in Drug Discovery & Development, 2, 96 (1999); (c) Cohen, P., Current Opinion in Chemical Biology, 3, 459 (1999); (d) Garcia-Echeverria et. al., Med. Res. Rev, 20, 28 (2000); (e) Blume-Jensen et al., Nature, 411, 355 (2001) and references therein; (f) Druker et. al., Nature Medicine, 2, 561 (1996); (g) Taylor et al., Current Opinion in Chemical Biology, 1, 2219 (1997); (h) Schindler et. al., Science, 289, 1938 (2000).) As each scaffold presents unique opportunities for the presentation of functional groups to the kinase active site, there is a need for efficient and flexible methods for preparing libraries of each of these inhibitor classes. The present invention fulfills these and other needs.